The ability to reconstitute adult skin with functional skin appendages has long been a major clinical objective for dermatologists and surgeons because reconstituted skin can be used to treat skin wounds or disorders with reduced scarring and enhanced cosmetic appeal (Lee et al., Tissue Engineering: Part C, 17:(4) 391-400 (2011)). Skin damage and loss can occur for many different reasons, such as genetic disorders, chronic wounding or acute trauma such as burns. The damage can be substantial with no possibility of skin regeneration. Burn wounds in particular can be deep and extensive and can often be fatal in the absence of treatment. Currently, the most common treatment method for burn victims includes surgery to facilitate wound closure, followed by autologous skin grafting, where epidermis from an undamaged site is harvested and applied to the full-thickness wound. However, there are significant limitations to these procedures and the repair and management of full-thickness skin damage resulting from burns is still a significant clinical challenge (Shevchenko et al., J Royal Soc, 7:229 (2009); Juhasz et al., Derm Res Prac, 2010:210150 (2010)).
Tissue engineering to treat hair loss involves transplanting tissue grafts or cells into the target area to induce hair follicle formation. Hair follicle induction and growth requires active and continuous epithelial and mesenchymal interactions (Stenn and Paus, Physiol Reviews, 81:449-494, (2001)). However, not all cells obtained from hair follicle grafts are capable of inducing new hair follicle formation.
Previous studies have shown that isolated hair follicles from human scalp can be grafted onto the backs of immunodeficient mice and subsequently successfully return to normal cycling (Hashimoto et al., J Invest Derm 115:200 (2000)). Human hair follicle formation by grafting a skin substitute incorporated with cultured mutant (TSCII deficient) fibroblast cells has been reported (Li et al., Nat Comm, 2:235 (2010)). However, the regenerated hair follicle did not produce outgrowing hair shafts, which is a central element of a mature hair follicle (Chuong et al., J Invest Derm, 127:2098 (2007)). Others have shown that skin substitutes consisting of epithelial cells and modified mesenchymal cells can be used as transplantable grafts capable of inducing hair follicles (WO2011/160055A2).
US Patent application No. 20120095445 by Zheng, et al., describes methods and reagents for creating hair follicles in vitro from dissociated epidermal and dermal cells, which are cultured and passaged separately, then combined to form hair follicles which are subsequently implanted. Sonic hedgehog pathway agonists are used to increase the trichogenicity of the dissociated cells. This process is limited, however, since the hair follicles must be formed in vitro, then implanted.
US Patent Publication No. 2011/0321180 to Lee et al. discloses the formation of murine hair follicles from freshly isolated murine multipotential precursor cells from neonatal mice. Because the number of freshly isolated cells is limited, there is a need for regenerating skin using culture-expanded cells. Unfortunately, culturing pluripotent cells often results in a loss in the ability of the cells to differentiate into desired cell types.
Therefore, it is an object of the invention to provide compositions and methods for producing reconstituted skin and methods of use thereof where dissociated cells are implanted directly into the site where hair is desired, and hair follicles form in situ.